ABSTRACT In type I diabetes, pancreatic islets are destroyed by an autoimmune reaction. Thus, a general strategic goal for preventing islet loss is to locally suppress this autoimmune response, while avoiding systemic immune suppression. FoxP3-expressing T regulatory cells (Tregs) are immune cells that play a central role in local tolerance, and therefore could serve as a potential platform for a cell therapy to prevent islet loss. Tregs have many attractive attributes as a suppressive cell therapy: they can exert dominant immunosuppressive actions, through cell-cell interactions or production of suppressive cytokines; they can in principle be long lived; and tolerance induced by Tregs can persist through infectious tolerance by conferring tolerogenic properties to neighboring cells. In mouse models, single infusion of Tregs can prevent and reverse diabetes indefinitely. The encouraging preliminary data in mouse models have let to the development of ongoing clinical trials of Treg cell therapies in humans. Nonetheless, Treg cell therapy presents several key challenges: it is hard to specifically target them to the islets and avoid systemic suppression, they are hard to expand in sufficient numbers (especially relative to conventional effector T cells), and their cell fate can be changed, especially in particular inflammatory microenvironments. In the past few years, however, there have been remarkable advances in using conventional T cells for cancer therapy (e.g. CAR T cells), including an explosion of new synthetic biology tools that can be used to precisely target T cells to disease tissues, to control their fate and proliferation, and to even give them the capability to locally deliver non-natural therapeutic payloads. Here our goal is to bring these new tools to bear on the problem of engineering improved islet-specific therapeutic suppressive cells. Our aims are to: 1) engineer natural Tregs with improved targeting and recognition of islets 2) develop tools to selectively expand therapeutic Tregs in vivo and enhance their stability 3) design synthetic suppressor cells that disarm effectors, dampen inflammation and promote islet repair These approaches wed cutting-edge synthetic biology with multiple strategies for developing therapeutic suppressor cells. We will establish proof-of-concept data using mouse model of autoimmune diabetes and apply them to test engineered human Tregs targeted to human islets using a humanized mouse model of autoimmune islet inflammation. Successful completion of this study will provide preclinical data for future implementation of these strategies in clinical trials.